Assessing An Alternative Method Of Storing Mosquito Blood Meals For Human DNA Profiling

Abstract

A simple method for storing mosquito blood meal samples which permits extraction and characterization of human DNA after polymerase chain reaction (PCR) amplification of target DNA sequences was tested. Abdomens of blood-fed and partially fed An. gambiae s.l, An. funestus and Cx. quinquefasciatus mosquitoes from an area of intense malaria transmission in western Kenya were directly expressed onto filter paper, air dried and stored at both room and refrigerated temperatures. These were compared with field collected samples stored in liquid nitrogen. Some samples were stored for2 weeks while others for 8 weeks. DNA was extracted using either the Chelex or Isoquick nucleic acid extraction protocols and amplified at human hypervariable loci TCII, VWA and DIS80. The amplified products were separated by polyacrylamide gel electrophoresis and visualized by silver staining. In preliminary experiments, a much higher success rate in amplification was observed 111 samples extracted usmg Isoquick rather than Chelex (P=O.OOO1); Isoquick was therefore used in subsequent experiments. The results showed that blooded abdomens stored on dried filter papers could be amplified with greater than 80% success for any locus, storage temperature, mosquito species, or storage duration. Some significant, but relatively minor differences were observed: blood meals from An. gambiae amplified at a slightly higher rate than those of An. funestus and Cx. quinquefasciatus (P=O.006), loci TCll and VWA amplified somewhat more successfully than DIS80 (P=0.0003), and storage temperature had small but significant effect (P=O.Ol)..on amplification success. No effect of storage duration was observed, indicating that the DNA remain.ed viable and could be extracted and typed from filter paper blotted mosquito blood meals, dried and stored for up to 8 weeks. These results show that filter papers which are cheap, readily available, easy to transport and store, work well compared to cryopreservation and refrigeration, when used to store mosquito blood meals for human DNA profiling studies.